CRISPR/Cas9-based functional analysis of yellow gene in the diamondback moth, Plutella xylostella

The diamondback moth,Plutella xylostella(L.), is an economically important pest of cruciferous crops worldwide. This pest is notorious for rapid evolution of the resistance to different classes of insecticides, making it increasingly difficult to control. Genetics-based control approaches, through manipulation of target genes, have been reported as promising supplements or alternatives to traditional methods of pest management. Here we identified a gene of pigmentation (yellow) inP. xylostella,Pxyellow, which encodes 1674 bp complementary DNA sequence with four exons and three introns. Using the clustered regularly interspersed palindromic repeats (CRISPR)/CRISPR-associated protein 9 system, we knocked outPxyellow, targeting two sites in Exon III, to generate 272 chimeric mutants (57% of the CRISPR-treated individuals) with color-changed phenotypes of the 1st to 3rd instar larvae, pupae, and adults, indicating thatPxyellowplays an essential role in the body pigmentation ofP. xylostella. Fitness analysis revealed no significant difference in the oviposition of adults, the hatchability of eggs, and the weight of pupae between homozygous mutants and wildtypes, suggesting thatPxyellowis not directly involved in regulation of growth, development, or reproduction. This work advances our understanding of the genetic and insect science molecular basis for body pigmentation ofP. xylostella, and opens a wide avenue for development of the genetically based pest control techniques usingPxyellowas a screening marker.